5'-Nucleotides, inter alia, 5'-IMP and 5'-GMP have a flavor enhancing activity and are widely used as a seasoning agent. As the method for producing 5'-nucleotides, there are known a method for enzymatically decomposing RNA present in yeast cells (Journal of Agricultural and Chemical Society of Japan, 34, 489, 1969), a method for culturing a microorganism having an ability to produce 5'-IMP Agricultural and Biological Chemistry, 46, 2557 (1982)!, a method for chemical phosphorylation of nucleosides such as inosine and guanosine Bulletin of the Chemical Society of Japan, 42, 3505-3508 (1969)!, and the like. Currently, it is considered that the chemical phosphorylation utilizing these nucleosides would be the most advantageous from an industrial standpoint, because inosine and guanosine are easily obtained by fermentation. However, in the case of performing chemical phosphorylation, large quantities of chlorides are used at a lower temperature Bulletin of the Chemical Society of Japan, 42, 3505-3508 (1969)! so that the method is not desired from viewpoints of both costs and environmental hygiene. Accordingly, enzymatic phosphorylation of nucleosides under mild conditions is desired. Hitherto, biochemical phosphorylation of purine nucleosides using cultured cells of various microorganisms has been reported (Japanese Published Unexamined Patent Application Nos. 116698/83 and 230094/88) but is inferior to chemical phosphorylation in terms of conversion rate and yield.
As an inosine-phosphorylating enzyme, inosine kinase (EC 2.7.1.73) is known but the crude active fraction derived from animal tissues or microorganisms is only reported The Enzymes, Vol. IX, 54-56 (1973), Academic Press; Nucleosides and Nucleobases in Microorganisms, pp. 66, (1983), Academic Press!. The presence of a guanosine kinase activity which phosphorylates guanosine in E. coli is also suggested J. Gen. Microbiol., 135, 1263-1273 (1989)!. However, the activity is low and it is extremely difficult to purify the enzyme so that it is far beyond industrial utilization. In addition, physicochemical properties of the enzyme are not clarified. In any event, inosine-guanosine kinase is an enzyme that has been neither isolated nor purified.
An object of the present invention is to provide an inosine-guanosine kinase and also to provide a method for preparing 5'-nucleotides, inter alia, 5'-IMP and 5'-GMP, which have been widely utilized as a seasoning agent, industrially at low costs.